Cheeseman et al., p.1
Supplementary Material
Supplementary Table 1. Worm Strains used in this study.
Strain # / GenotypeTH32 / unc-119(ed3) III; ruIs32[pAZ132; pie-1/GFP::histone H2B] III; ddIs6 [GFP::tbg-1; unc-119(+)]
OD1 / unc-119(ed3) III; ltIs1 [pIC22; pie-1/GFP::knl-3; unc-119 (+)]
OD8 / unc-119(ed3) III; ltIs4 [pIC32; pie-1/GFP-TEV-STag::mis-12; unc-119 (+)]
OD9 / unc-119(ed3) III; ltIs5 [pIC36; pie-1/GFP-TEV-STag::kbp-1; unc-119 (+)]
OD10 / unc-119(ed3) III; ltIs6 [pIC35; pie-1/GFP-TEV-STag::kbp-5; unc-119 (+)]
OD11 / unc-119(ed3) III; ltIs7 [pIC41; pie-1/GFP-TEV-STag::kbp-4; unc-119 (+)]/+
OD13 / unc-119(ed3) III; ltIs9 [pIC34; pie-1/GFP-TEV-STag::kbp-3; unc-119 (+)]/+
Supplementary Table 2. dsRNAs used in this study.
• dsRNAs were amplified from either N2 genomic DNA (61, 84, 93, and 94) or gene specific cDNAs (171, 199-204).
RNA# / Gene # / Name / [ ] mg/ml / Oligo #1 / Oligo #2 / Template61 / C02F5.1 / knl-1 / 1.9 / TAATACGACTCACTATAGGccgctgaaatggatacgagt / AATTAACCCTCACTAAAGGccatgctaatgtcttcacacg / N2 genomic DNA
84 / W01B6.9 / ndc-80 / 1.6 / AATTAACCCTCACTAAAGGccccagtctgagtcaacctc / TAATACGACTCACTATAGGccaactcgctttgaatttcc / N2 genomic DNA
93 / T03F1.9 / hcp-4 / 1.4 / aattaaccctcactaaaggggaaatgtacggagcgaaaa / taatacgactcactataggacattgttggtgggtccaat / N2 genomic DNA
94 / F58A4.3 / hcp-3 / 1.7 / AATTAACCCTCACTAAAGGgccgatgacaccccaattat / TAATACGACTCACTATAGGgttccttccggctctcatc / yk511e10
171 / T10B5.6 / knl-3 / 1.2 / AATTAACCCTCACTAAAGGatgtctcaaaaatcaaacgacacc / TAATACGACTCACTATAGGgtcgagaaaacttccgtgaag / yk494e5
199 / Y47G6A.24 / mis-12 / 2.6 / AATTAACCCTCACTAAAGGatggagagcgaattgtcggga / TAATACGACTCACTATAGGggttaaactggagacattcgtg / yk156d5
200 / Y92C3B.1 / kbp-4 / 3.5 / AATTAACCCTCACTAAAGGaaaaatgcaaaaatcaatcgac / TAATACGACTCACTATAGGgaaaatttcctcggcgacct / yk628f9
201 / F26F4.13 / kbp-2 / 2.6 / AATTAACCCTCACTAAAGGtaaagctgcagtggatgctg / TAATACGACTCACTATAGGgcttttgctgttgtgactcg / N2 cDNA
202 / F26H11.1 / kbp-3 / 2.4 / AATTAACCCTCACTAAAGGtttttcggcaaaatggagtc / TAATACGACTCACTATAGGttgatacgtggatgaaaacca / N2 cDNA
203 / C34B2.2 / kbp-5 / 2.3 / AATTAACCCTCACTAAAGGaaatgaatccaaatcataatgaagg / TAATACGACTCACTATAGGagagaatcacatcatgaggaaatatg / N2 cDNA
204 / R13F6.1 / kbp-1 / 3.4 / AATTAACCCTCACTAAAGGtgtcgaaggaagaacgaaaga / TAATACGACTCACTATAGGgctgccacggtattgacttc / yk732e6
Supplementary Table 3. Summary of supplemental movies.
• In all videos, the anterior of the embryo is to the left and the posterior of the embryo is to the right.
• All images were acquired using widefield deconvolution microscopy
• For TH32 (GFP-Histone H2B + GFP--tubulin), one 3D z-stack (6 images at 2 µm steps) was acquired every 10s
• For GFP-kinetochore protein strains, one 3D z-stack (6 images at 1 µm steps) was acquired every 10s
• Maximal intensity projections of z-stacks were used to generate each movie frame
• All images from the same strain were scaled with respect to the wild type control
• Video playback is at 10 frames/sec
Video #
/ Strain / Fluorescent Markers / RNAi1 / TH32 / GFP-Histone H2B; GFP--tubulin / None (wild type)
2 / TH32 / GFP-Histone H2B; GFP--tubulin / knl-1 (#61)
3 / TH32 / GFP-Histone H2B; GFP--tubulin / knl-3 (#171)
4 / TH32 / GFP-Histone H2B; GFP--tubulin / ndc-80 (#84)
5 / TH32 / GFP-Histone H2B; GFP--tubulin / kbp-3 (#202)
6 / TH32 / GFP-Histone H2B; GFP--tubulin / mis-12 (#199)
7 / TH32 / GFP-Histone H2B; GFP--tubulin / kbp-1 (#204)
8 / TH32 / GFP-Histone H2B; GFP--tubulin / kbp-2 (#201)
9 / TH32 / GFP-Histone H2B; GFP--tubulin / kbp-4 (#200)
10 / OD1 / GFP-KNL-3 / None (wild type)
11 / OD1 / GFP-KNL-3 / mis-12 (#199)
12 / OD1 / GFP-KNL-3 / kbp-1 (#204)
13 / OD8 / GFP-MIS-12 / None (wild type)
14 / OD8 / GFP-MIS-12 / mis-12 (#199)
15 / OD9 / GFP-KBP-1 / None (wild type)
16 / OD10 / GFP-KBP-5 / None (wild type)
17 / OD10 / GFP-KBP-5 / mis-12 (#199)
18 / OD11 / GFP-KBP-4 / None (wild type)
19 / OD11 / GFP-KBP-4 / mis-12 (#199)
20 / OD13 / GFP-Spc25KBP-3 / None (wild type)
21 / OD13 / GFP-Spc25KBP-3 / mis-12 (#199)
22 / OD13 / GFP-Spc25KBP-3 / kbp-1 (#204)
23 / OD1 / GFP-KNL-3 / knl-3 (#171)
24 / OD1 / GFP-KNL-3 / knl-1 (#61)
25 / OD1 / GFP-KNL-3 / ndc-80 (#84)
26 / OD8 / GFP-MIS-12 / knl-3 (#171)
27 / OD8 / GFP-MIS-12 / knl-1 (#61)
28 / OD8 / GFP-MIS-12 / ndc-80 (#84)
29 / OD9 / GFP-KBP-1 / knl-3 (#171)
30 / OD9 / GFP-KBP-1 / knl-1 (#61)
31 / OD9 / GFP-KBP-1 / ndc-80 (#84)
32 / OD10 / GFP-KBP-5 / knl-3 (#171)
33 / OD10 / GFP-KBP-5 / knl-1 (#61)
34 / OD10 / GFP-KBP-5 / ndc-80 (#84)
35 / OD11 / GFP-KBP-4 / knl-3 (#171)
36 / OD11 / GFP-KBP-4 / knl-1 (#61)
37 / OD11 / GFP-KBP-4 / ndc-80 (#84)
38 / OD13 / GFP-Spc25KBP-3 / knl-3 (#171)
39 / OD13 / GFP-Spc25KBP-3 / knl-1 (#61)
40 / OD13 / GFP-Spc25KBP-3 / ndc-80 (#84)